Abstract

We studied the changes in the higher-order structure of a megabase-size DNA (S120-1 DNA) under different spermidine (SPD) concentrations through single-molecule observations using fluorescence microscopy (FM) and atomic force microscopy(AFM). We examined the difference between the folding transitions in S120-1 DNA and sub-megabase-size DNA, T4 DNA (166 kbp). From FM observations, it is found that S120-1 DNA exhibits intra-chain segregation as the intermediate state of transition, in contrast to the all-or-none nature of the transition on T4 DNA. Large S120-1 DNA exhibits a folding transition at lower concentrations of SPD than T4 DNA.AFM observations showed that DNA segments become aligned in parallel on a two-dimensional surface as the SPD concentration increases and that highly intense parallel alignment is achieved just before the compaction. S120-1 DNA requires one-tenth the SPD concentration as that required by T4 DNA to achieve the same degree of parallel ordering. We theoretically discuss the cause of the parallel ordering near the transition into a fully compact state on a two-dimensional surface, and argue that such parallel ordering disappears in bulk solution.

Received 22 August 2011Accepted 17 November 2011Published online 08 December 2011

Acknowledgments:

This work was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan (18GS0421) and from the Japan Society for the Promotion of Science (22510123, 23240044, and 23657068). Y. S. is a recipient of the JSPS Research Fellow.

Article outline:I. INTRODUCTIONII. EXPERIMENTAL METHODSA. MaterialsB. Preparation of genomicDNA from R. appendicifer strain 120-1TC. Direct observation of the change in the conformation of DNA in bulk solution and on a glass surface by FMD. AFM observation of single DNA moleculesIII. RESULTSA. FM observations of the higher-order structure of single DNA moleculesB. AFM observation of the higher-order structure of single DNA moleculesIV. DISCUSSIONA. Evaluation of the degree of parallel alignmentB. Enhanced parallel ordering near compaction on a 2D surfaceC. Difference between 2D surface and 3D solutionD. Difference in the higher-order structural change between S120-1 DNA and T4 DNAE. Intra- and inter-chain segregation

Abstract

We studied the changes in the higher-order structure of a megabase-size DNA (S120-1 DNA) under different spermidine (SPD) concentrations through single-molecule observations using fluorescence microscopy (FM) and atomic force microscopy(AFM). We examined the difference between the folding transitions in S120-1 DNA and sub-megabase-size DNA, T4 DNA (166 kbp). From FM observations, it is found that S120-1 DNA exhibits intra-chain segregation as the intermediate state of transition, in contrast to the all-or-none nature of the transition on T4 DNA. Large S120-1 DNA exhibits a folding transition at lower concentrations of SPD than T4 DNA.AFM observations showed that DNA segments become aligned in parallel on a two-dimensional surface as the SPD concentration increases and that highly intense parallel alignment is achieved just before the compaction. S120-1 DNA requires one-tenth the SPD concentration as that required by T4 DNA to achieve the same degree of parallel ordering. We theoretically discuss the cause of the parallel ordering near the transition into a fully compact state on a two-dimensional surface, and argue that such parallel ordering disappears in bulk solution.

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Critical behavior of megabase-size DNA toward the transition into a compact state